Splenocyte proliferation assay
One week after the last immunization, spleens of 3 mice in each group
were collected and disrupted with the plunger of a syringe. After
straining through a 100 µm mesh, red blood cells were lysed in
erythrocyte lysis buffer (Solarbio, Beijing, China). Splenocytes
(2.5×105) were plated together with the purified
ZIKV-E antigen (10 μg/ml) in 96-well plates. After incubation for 44 h,
10 µl/well CCK-8 (KeyGEN Biotech, Nanjing, China) was added to the cells
to determine the cell proliferation in the next step. After cultivation
for another 4 h, the absorbance was measured at 450 nm using a
microplate reader (Thermo Fisher Scientific, Waltham, MA, USA). The
proliferation index (PI) was calculated as: (ODstimulated cultures − ODnon-stimulated
cultures)/(ODnon-stimulated cultures −
ODcontrol cultures).